This application describes a 5-year program designed to provide training for Dr. Michael Sebert in the skills necessary to become an independent researcher in the field of microbial pathogenesis. Dr. Sebert is currently completing his fellowship training in pediatric infectious diseases. His present work in the laboratory of Dr. Jeffrey Weiser, his sponsor for this application, has provided preliminary exposure to the study of the molecular mechanisms of bacterial infection. The educational plan outlined herein consists of both a comprehensive didactic program intended to provide a solid foundation in molecular genetics, prokaryotic cell biology, immunology, and bacterial pathogenesis as well as a roadmap for the in-depth laboratory experience necessary for his maturation into a productive, independent investigator. Colonization of the upper respiratory tract by the major bacterial pathogen Streptococcus pneumoniae serves as the common first step in the pathogenesis of the wide range of disease processes caused by this organism. How S. pneumoniae senses and adapts to this environment is unknown. Genomic analyses of the pneumococcus have revealed over a dozen putative two-component signal transduction systems. PreliminaryInvestigations have revealed that one of these systems, CiaRH, is required for persistence of the organism in the infant rat nasopharynx and that a portion, but not all, of this failure to colonize effectively can be attributed to downregulation of a putative serine protease, HtrA. The hypothesis to be tested in the proposed research is that a set of genes is regulated by the CiaRH system that collectively account for the requirement of this system for effective carriage. The first specific aim toward this objective is the biochemical and molecular characterization of the HtrA protein in order to define its precise role in nasal carriage. The second specific aim of this project is the definition of the sequence elements upstream of htrA responsible for its regulation by CiaRH and the subsequent usage of this information to identify other genes under the control of CiaRH that contribute to colonization. Together these lines of investigation hold the potential to expand the understanding of the molecular basis of this critical step in the pathogenesis of S. pneumoniae.